Polymerase chain reaction-based method for the identification of leishmania (viannia) Braziliensis and leishmania (viannia) guyanensis in mucosal tissues conserved in paraffin

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Abstract

Introduction: In the Americas, mucosal leishmaniasis is primarily associated with infection by Leishmania (Viannia) braziliensis. However, Leishmania (Viannia) guyanensis is another important cause of this disease in the Brazilian Amazon. In this study, we aimed at detecting Leishmania deoxyribonucleic acid (DNA) within paraffi n-embedded fragments of mucosal tissues, and characterizing the infecting parasite species. Methods: We evaluated samples collected from 114 patients treated at a reference center in the Brazilian Amazon by polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) analyses. Results: Direct examination of biopsy imprints detected parasites in 10 of the 114 samples, while evaluation of hematoxylin and eosin-stained slides detected amastigotes in an additional 17 samples. Meanwhile, 31/114 samples (27.2%) were positive for Leishmania spp. kinetoplast deoxyribonucleic acid (kDNA) by PCR analysis. Of these, 17 (54.8%) yielded amplifi cation of the mini-exon PCR target, thereby allowing for PCR-RFLP-based identifi cation. Six of the samples were identifi ed as L. (V.) braziliensis, while the remaining 11 were identifi ed as L. (V.) guyanensis. Conclusions: The results of this study demonstrate the feasibility of applying molecular techniques for the diagnosis of human parasites within paraffi nembedded tissues. Moreover, our fi ndings confi rm that L. (V.) guyanensis is a relevant causative agent of mucosal leishmaniasis in the Brazilian Amazon.

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Prestes, S. R., Guerra, J. A. de O., Romero, G. A. S., Magalhaes, L. K. C., Santana, R. A. G., Maciel, M. G., … Silveira, H. (2015). Polymerase chain reaction-based method for the identification of leishmania (viannia) Braziliensis and leishmania (viannia) guyanensis in mucosal tissues conserved in paraffin. Revista Da Sociedade Brasileira de Medicina Tropical, 48(5), 555–559. https://doi.org/10.1590/0037-8682-0132-2015

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